Stabilized Carbon‐Centered Radical‐Mediated Carbosulfenylation of Styrenes: Modular Synthesis of Sulfur‐Containing Glycine and Peptide Derivatives

Abstract Sulfur‐containing amino acids and peptides play critical roles in organisms. Thiol‐ene reactions between the thiol residues of L‐cysteine and the alkenyl fragments in the designed coupling partners serve as primary tools for constructing C─S bonds in the synthesis of unnatural sulfur‐containing amino acid derivatives. These reactions are favored due to the preference for hydrogen transfer from thiol to β‐sulfanyl carbon radical intermediates. In this paper, the study proposes utilizing carbon‐centered radicals stabilized by the capto‐dative effect, generated under photocatalytic conditions from N–aryl glycine derivatives. The aim is to compete with the thiol hydrogen, enabling radical C─C bond formation with β‐sulfanyl carbon radicals. This protocol is robust in the presence of air and water, offers significant potential as a modular and efficient platform for synthesizing sulfur‐containing amino acids and modifying peptides, particularly with abundant disulfides and styrenes.


Introduction
Sulfur-containing amino acids and their derivatives play crucial roles in biological processes.For example, S-adenosylmethionine serves as the methyl source for nearly all mammalian transmethylases, and most eukaryotic proteins originate from methionine (Scheme 1). [1]However, the bioprocesses involved in the formation of C─S bond during the synthesis of sulfurcontaining molecules in living organisms remain unexplored.The laboratory synthesis of sulfur-containing unnatural amino acids (UAAs) and peptides heavily relies on the utilization of thiol residues from naturally occurring L-cysteine and its derivatives. [2]mong the reported methods, thiol-ene reactions, a well-known DOI: 10.1002/advs.202402428type of "click reactions", are commonly used for constructing C─S bonds.Thiol-ene reactions have found extensive applications in synthetic methodologies, biofunctionalization, and polymer science, as reported. [3]he mechanism behind these reactions has been extensively studied and is well established.It involves the addition of thiyl radicals to C─C double bonds, generated from thiols in the presence of light or radical initiators, followed by hydrogen atom transfer (HAT) from another thiol molecule to the carbon radical.(Scheme 2A). [4]Despite the potential of difunctionalization of alkenes to construct molecular complexity in a single operation, which has attracted wide interest, [5] challenges persist in synthesizing sulfur-containing UAAs through the incorporation of carbosulfenylation of alkenes with thiols under mild and atmospheric conditions: 1) Thiols serve as effective hydrogen donors with bond dissociation energy of ≈87 and 80 kcal mol −1 for RS-H and ArS-H, respectively.The transfer of hydrogen to the -sulfanyl carboncentered radical intermediates from the thiols is favored due to the particularly low activation energy of the thermoneutral reaction (E e0 ). [6]In fact, the facilitated hydrogen transfer from thiols to carbon radicals is the predominant pathway for repairing damage to nucleic acids and proteins caused by other free radicals or toxic species in biological systems. [7]2) The synthesis and modification of amino acids and peptide derivatives typically require polar solvents as the reaction medium due to their limited solubility in non-polar solvent.Hydrogen transfer from thiols to carbon radicals can be significantly accelerated by utilizing polar solvents because they stabilize the polar transition state. [8]3) When the reaction system is exposed to atmospheric oxygen, thiol-olefin cooxidation (TOCO) becomes the primary reaction pathway. [9]As a result, achieving mild and general carbosulfenylation of alkenes through the addition of thiyl radicals followed by radical-type C─C cross-coupling remains challenging (for a highly substrate depended photo-induced carbosulfenylation of activated styrenes (arylidenemalononitrile) via radical-polar cross-over process, see ref. [10]) From a broader perspective, investigations into the intermolecular carbosulfenylation of alkenes pose significant challenges, whether through the thiyl radical intermediates or twoelectron chemistry (Scheme 2B).Trost et al. pioneered carbosulfenylation by the electrophilic activation of alkenes with a sulfenylating agent, followed by the addition of alkynylalaneates, prepared through reactions involving alkynyl lithium reagents and triethylaluminum. [11]We recently achieved the arylation of thiiranium ion intermediates, leveraging the covalent nature of organozinc reagents. [12]Zhao et al. reported the enantioselective sulfenoarylation of N-allyl sulfonamides by reacting thiiranium ions with electron-rich phenols.This reaction was conducted in the presence of an indene-based chiral selenide catalyst and a Tf 2 NH additive (see ref. [13], for intramolecular carbosulfenylation of alkenes intermediated by thirranium ions). [14]Engle et al. [15] and Wang et al. [16] independently established creative sulfenoalkylation/sulfenoarylations of alkenes via metallacycle intermediates by pre-installing directing groups in alkene substrates and using organometallic agents (dialkylzincs and aryl boronic acids) as carbon sources.These reactions involve air-or moisture-sensitive thiiranium ions or metal complexes and primarily utilize organometallic reagents as carbon nucleophilic partners, thus limiting the reaction conditions.Inspired by the innovative studies on photocatalysis-enabled thiolene reactions, [17] Hao et al. recently developed an elegant carbosulfenylation of 1,3-butadiene through synergistic catalysis with iridium and titanium. [18]In this process, the formation of stabilized allyl radical intermediates was employed to inhibit the HAT process from thiol, [19] thereby enabling subsequent C─C bond coupling, which, turned out to be the limitation of the reaction with only 1,3-butadiene substrate.A more general carbosulfenylation protocol, allowing the modular incorporation of abundant sulfur compounds into amino acid and peptide derivatives under mild atmospheric conditions, is highly desirable.Photocatalyzed C(sp 3 )-H alkylations of glycinate derivatives have emerged as effective methods for preparing UAAs through various radical transformations, facilitating the modification of naturally occurring amino acid derivatives. [20]We realized that integrating thiyl radicals and carbon radical intermediates, derived from disulfides and glycine derivatives, with styrenes under photocatalyzed conditions (Scheme 2C) could overcome the above-mentioned challenges and provide the following advantages for carbosulfenylation: 1) A significantly large rate constant (≈1 × 10 4 times) for the inverse reaction of HAT from thiols to benzylic carbon radicals would effectively inhibit the thiol-ene side reaction. [6,21]2) The low reactivity of disulfides with the sulfanyl carbon-centered radical intermediates would prevent the formation of disulfenylation byproducts. [22]3) The capto-dative stabilization (or mero-stabilization) effect from two mutual substituents (-donor and -acceptor) in the radical intermediates of glycine derivatives, combined with the influence of high dielectric constant solvents-known to significantly enhance the capto-dative effect [23] and favored for peptide reaction-provides the opportune stability and reactivity of the glycinate-derived radical intermediates [24a] for radical-type C─C bond formation.4) The suitable reactivity of -sulfanyl carbon radicals in the presence of stabilized glycinate carbon radicals would circumvent the influences of atmospheric oxygen (TOCO reaction) and water additives, enabling the reaction to be conducted with protonic functionalities in a range of polar solvents without requiring an inert atmosphere.This would enable the modular synthesis of a variety of sulfur-containing UAAs and peptide derivatives.Herein, we present our studies on the photocatalyzed carbosulfenylation of styrenes with disulfides and glycine derivatives under ambient conditions, facilitated by radical-type C─C bond formation enabled by the capto-dative stabilization effect.

Results and Discussion
We initially investigated the catalytic system with Nphenylglycine ethyl ester 1, 1,1-diphenylethylene 2 and diphenyldisulfide 3 in acetonitrile (MeCN) as a model reaction.The system was irradiated with a 15 W blue light (462 nm) using light emitting diode (LED) under an air atmosphere (Table 1).
Upon employing Ir(ppy) 3 as photocatalyst, sulfanyl glycine derivative 4 was isolated in a 61% yield (entry 1).Encouraged by these results, alternative photocatalysts were screened.Both [Ir(ppy) 2 (dtbbpy)]PF 6 (entry 2) and (Ir[dF(CF 3 )ppy] 2 (dtbbpy))PF 6 (entry 3) effectively promoted the reaction, yielding 4 in 69% and 68% isolated yields, respectively.Other photocatalysts such as ruthenium-based complexes and organic dyes, proved much less effective (entry 4 and Table S1, Supporting Information).Throughout the screening process, hydrosulfenylation of 2 was detected as a minor reaction.We hypothesized that the in situ generated thiophenol was responsible for this due to its hydrogen content.Therefore, we attempted to use a base as an additive for the deprotonation process; [19a] however, this did not lead to an increase in the yield (entries 5 and 6).Substituting the disulfide reagent with N-(phenylthio)phthalimide, a typical [N-S] agent, resulted in a low yield (entry 7).Additionally, control experiments illustrated that the reaction proceeded smoothly under nitrogen (entry 8), confirming the essential role of both light and the photocatalyst in the production of the anticipated product (Table S8, Supporting Information).The regioselectivity of the carbosulfenylation of styrene was confirmed through single-crystal X-ray crystallographic analysis of 4 (Figure S20, Supporting Information).As the reaction proceeded efficiently in air, we investigated its compatibility with water.Using MeCN and water (4/1, v/v) as cosolvents resulted in a satisfactory yield of the sulfanyl derivative of glycinate (entry 9), suggesting the potential of this protocol for applications under biologically compatible conditions and transformations.
in Scheme 4. The influence of the electron density of the Naryl moiety on the protocol's outcome was tested (37-41), revealing that electron-rich phenyl rings led to slightly low yields (38 and 41).The heteroaryl functionality of glycinate was successfully introduced using this procedure (42).Both phenol-and alkyl alcohol-derived glycinates were compatible (43-45).Additionally, N-arylaminoacetonitrile could be utilized in this reaction (46).The satisfactory performance of amide derivatives of glycine (47 and 48), along with their excellent functional group tolerance, prompted us to explore the selective sulfenylative alkylation of various peptides.We prepared a series of dipeptides from N-aryl glycine with leucine, phenylalanine, methionine, and tyrosine, subjecting them to standard conditions.Sulfenylative dipeptides were successfully produced in moderate yields (49-53).Additionally, tripeptides and tetrapeptides underwent carbosulfenylation smoothly (54 and 55).Drug molecules, including podophyllotoxin-an antineoplastic glucoside and antitumor agent-and linezolid-an antibacterial, were coupled with N-  phenyl glycine and assessed under standard conditions, leading to the isolation of the corresponding products in moderate yields (56 and 57).
Encouraged by the method's generality and its resistance to oxygen and water, we extended the photochemical protocol to the carbosulfonylation [25] of styrenes, exploring the scope of glycinates and thiosulfonates (R 1 SO 2 SR 2 ). [26]By slightly modifying the reaction conditions and replacing MeCN with DMF (N,Ndimethylformamide), we achieved the modular synthesis of a range of sulfonyl glycinate derivatives, as shown in Scheme 5. Aryl sulfonyl functionalities, whether bearing electron-pushing or electron-withdrawing, substituents were well tolerated (58-62).S-phenyl alkanethiosulfonates with both acyclic (methyl and butyl) and cyclic (cyclopropyl) carbon chains were successfully transferred (63-65).Building on the success of the carbosulfenylation, this analogous reaction proved compatible with various styrenes (66-68) and glycinate derivatives (69-71), yielding sulfonylated products in good to excellent yields.
To demonstrate the practicality of this protocol in synthesizing sulfur-containing UAAs, a gram-scale reaction was conducted, yielding product 58 in 61% yield (Scheme 6A).The N-PMP (4-methoxyphenyl) group was easily removed to obtain the amino ester intermediate, which was then amidated with Boc-Gly (N-(tert-butoxycarbonyl)glycine), resulting in an overall yield of 54% (Scheme 6B).Reduction of the ester group with lithium aluminum hydride (LiAlH 4 ) afforded the -amino alcohol 74 (Scheme 6C).In addition, the broad scope of glycine derivatives, peptides, disulfides, and styrenes prompted us to test the compatibility of the carbosulfenylation reaction with biomolecules under ambient conditions.The reactions conducted in MeCN:H 2 O (4:1, v/v) using various biomolecules as additives, including nucleobase, protein, amino acid, saccharide, and biotin showed little impact on the outcomes of the reactions (Table 2), further indicating the potential utility of this protocol for biological applications.Several control experiments were conducted to elucidate the carbosulfenylation mechanism (Scheme 7).The reaction was completely inhibited by the addition of an excess of the radical inhibitor 2,2,6,6-tetramethyl-1-peperidyloxy (TEMPO) and afforded the high-resolution mass spectrometry (HRMS)detectable species 75 and 76 (Scheme 7A), which were formed through the coupling of carbon radical intermediates with TEMPO.Under the optimized conditions of the model reaction, the homocoupling product of glycinate (79) was isolated as a byproduct in 13% yield (Scheme 7B, Equation ( 1)).These results indicate the involvement of both the carbon-centered radical of glycinate and the -sulfanyl carbon radical in this process.Additionally, a mixture of sulfide products 77 and 78 (1:1 ratio, as determined by proton nuclear magnetic resonance ( 1 H NMR)) was obtained.Control experiments were conducted with and without glycinate (Scheme 7B, Equations (2) and ( 3)), indicating that a photocatalyst was necessary for the generation of 77 and 78.( 19a,27] Disulfides are effective thiyl radical acceptors [28] ; therefore, the exchange of sulfides (3a and 3c, Scheme 7C) was observed under irradiation with blue light without a photocatalyst.This occurs because the homolytic cleavage of disulfides to produce thiyl radicals under light irradiation is possible, albeit inefficient [2a] in the addition of styrene.The absence of carbonsulfenylation product 4 when the imine analog 81 was used under standard conditions instead of glycinate 1 ruled out the possibility of sequential single-electron oxidation of glycinate (Scheme 7D).A series of Stern-Volmer fluorescence quenching experiments with substrates showed that only glycinate effectively quenched the excited state of the photocatalyst (Scheme 7E), whereas disulfide or thiosulfonate were relatively less likely to interact with it.These findings were consistent with the literature reports indicating that the standard reduction potentials of disulfides (e.g., 1,2-di-p-tolyldisulfide with E 1/2 red = −2.15V vs SCE [26] and 1,2-dibutyldisulfide with E 1/2 red = −2.14V vs SCE [29] ) do not match the value of the photocatalyst ([Ir(ppy) 2 (dtbbpy)](PF 6 ) with E 1/2 (PC •─ /PC) = −1.51V vs SCE).Light on/off experiments revealed that a continuous light irradiation was essential to achieve a satisfactory yield of the product (Figure S17, Supporting Information).
Based on the aforementioned results and a literature review, a plausible mechanism for the modular synthesis of sulfur-containing UAAs and peptide modification was proposed (Scheme 8).First, upon irradiation with blue light, the excited state of the photocatalyst [Ir(ppy) 2 (dtbbpy)](PF 6 )* is generated and undergoes a single-electron transfer event with the glycine derivative.This generates the radical cation intermediate A (precursor of the carbon radical B) and the radical anion PC •─ (glycine ester E 1/2 ox = +0.31V vs SCE; [20k] [Ir(ppy) 2 (dtbbpy)](PF 6 ) E 1/2 (PC*/PC •─ ) = +0.66V vs SCE).The thiyl radical, derived from the irradiation of disulfide, a process that can be accelerated by the presence of photocatalyst, plays dual roles: it oxidizes PC •─ to regenerate the photocatalyst (E 1/2 (PhS • /PhS ─ ) = +0.16V vs SCE) [30] and adds to the C─C double bond of styrene, forming the -sulfanyl radical intermediate C. The benzyl carbon radical C is then trapped by intermediate B to yield the desired product.Inhibiting thiol-ene (E) and TOCO (D) side reactions hinges on the preformed carbon radical B, which is stabilized by two mutual substituents and is an excellent carbon radical acceptor. [31]However, a pathway involving single-electron transfer from PC •─ to thiosulfonates in carbosulfonylation cannot be completely ruled out.In some cases, PC •─ may directly reduce thiosulfonates (e.g., S-(4-methylphenyl)-4-methylbenzenethiosulfonate: E p1 = −1.432V vs SCE [26] ).

Conclusion
In summary, a visible-light-induced carbosulfenylation of styrenes with N-aryl glycinates and disulfides was developed.This protocol allows the straightforward synthesis of sulfurcontaining amino acid and peptide derivatives by varying the reagent components.The strategic use of N-aryl glycinate substrates, which exhibit capto-dative effects, stabilizes carboncentered radicals and act as coupling partners for C─C bond formation.This feature renders the reaction system insensitive to air and moisture, while maintaining productivity in protic solvents.34][35][36][37][38][39][40][41][42][43][44]

Scheme 2 .
Scheme 2. Overview of state-of-the-art.A) Hydrogen atom transfer (HAT) in thiol-ene reactions and challenges in radical-type C─C bond formation; B) Carbosulfenylation of alkenes; C) Stabilized carbon radical-mediated carbosulfenylation of styrenes for modular synthesis of sulfur-containing amino acid and peptide derivatives.

Table 1 .
Optimization of reaction conditions.